Genome-wide association studies and fine-mapping identify genomic loci for n-3 and n-6 polyunsaturated fatty acids in Hispanic American and African American cohorts

Omega-3 (n-3) and omega-6 (n-6) polyunsaturated fatty acids (PUFAs) play critical roles in human health. Prior genome-wide association studies (GWAS) of n-3 and n-6 PUFAs in European Americans from the CHARGE Consortium have documented strong genetic signals in/near the FADS locus on chromosome 11. We performed a GWAS of four n-3 and four n-6 PUFAs in Hispanic American (n = 1454) and African American (n = 2278) participants from three CHARGE cohorts. Applying a genome-wide significance threshold of P < 5 × 10−8, we confirmed association of the FADS signal and found evidence of two additional signals (in DAGLA and BEST1) within 200 kb of the originally reported FADS signal. Outside of the FADS region, we identified novel signals for arachidonic acid (AA) in Hispanic Americans located in/near genes including TMX2, SLC29A2, ANKRD13D and POLD4, and spanning a > 9 Mb region on chromosome 11 (57.5 Mb ~ 67.1 Mb). Among these novel signals, we found associations unique to Hispanic Americans, including rs28364240, a POLD4 missense variant for AA that is common in CHARGE Hispanic Americans but absent in other race/ancestry groups. Our study sheds light on the genetics of PUFAs and the value of investigating complex trait genetics across diverse ancestry populations.


The text in the figures
) is blurry and difficult to read, both when printed out and on the computer. If possible, would recommend trying to make the text in these figures clearer. They are an excellent part of the paper, and I would have liked to read them in detail. 2. A diagram clearly demonstrating the flow of the study and which cohorts/participants were used at each stage would be helpful for the reader to follow along. (Even if is just a supplemental figure). 3. The Discussion section of the paper is overall well-written. However, there are times that it seemed the results were simply being restated and summarized. A deeper interpretation of these results, and the implications they have would greatly benefit this manuscript. This is something that could simply be elaborated on at the end of the Discussion. For example, the authors state: "Our findings provide new insight into the complex genetics of circulating PUFA levels that reflect, in part, their response to evolutionary pressures across the course of human history." With respect to human health and disease, what are the implications of these findings? It would also be useful to elaborate on future work. The authors state clearly that there is a continued need to expand GWAS studies in cohorts with genetic ancestry that is reflective of the general population. But aside from this, and given the results found in the present manuscript, what would be the next steps in this line of research? This is also something that could be elaborate on in the second to last paragraph of the Discussion section, or briefly mentioned in the last paragraph of the Discussion section. 4. The author's attempt at replication due to there being limited data for these traits is appreciated, but the replication scheme is difficult to understand. 5. Table 4 -please clarify which race group the discovery falls under and what race the replication results come from.

Reviewers' comments:
Reviewer #1 (Remarks to the Author): There are two major goals of the study. The first one is to examine whether the major loci identified in European Americans are shared across race/ancestry groups. The second one is to examine the evidence for genetic association unique to HIS and AFA populations.
Response: We thank the reviewer for the targeted synthesis of our manuscript and identification of points for improvement. Table S6, the EAF of rs174585 is 0.061 for AA but 0.275 for Hispanic. How to explain these differences?

Major comment: 1. The EAF between the discovery set and replication set is different a lot. For example in
Response: We agree that the EAF of some variants differ between the discovery and replication cohorts. The reason of the EAF differences is that we applied crossancestry replication analysis and some of the variants examined for replication show distinct allele frequencies across ancestry groups. We highlight the point about cross-ancestry differences in allele frequencies in the fourth paragraph of the section on replication/validation in the Results: "Some of the novel signals without crossancestry replication demonstrated large differences in allele frequencies across groups…" (p. 12).

This study performed cross-ancestry replication analysis in CHARGE and MESA, with validation using the multi-ancestry GWAS of lipids from the Global Lipids Genetics Consortium (GLGC). In the method section, I cannot easily follow this part.
Response: We thank the reviewer for this valuable comment. To improve clarity, we have now separated the 'Follow-up replication and validation analyses' paragraph into two paragraphs ('Cross-ancestry replication analysis' and 'Validation analysis') and added additional information to clarify the replication analysis and validation analysis (pp. 22-23).

In the Follow-Up Replication and Validation Analyses of the method section, this study seems to use summary statistics from different cohort studies. How do you compare the SNP signals found in different sample sizes and adjusted variables?
Response: We thank the reviewer for this insightful comment. We agree with the reviewer that it is important to check the consistency of covariate adjustments used in the discovery and replication/validation analyses.
The cross-ancestry replication analyses examining associations with fatty acid traits were performed by the CHARGE consortium investigators, including ourselves, and used covariate adjustments completely consistent with those presented in our discovery analyses. Specifically, these analyses were all adjusted for age, sex, study site (where appropriate) and PCs of ancestry.
For the validation analysis based on the lipid traits from GLGC resources, analyses were carried out separately by ancestry group at each cohort and residuals were generated separately in males and females adjusting for age, age 2 , PCs of ancestry and any necessary study-specific covariates (Graham et al.,PMID: 34887591). Thus, the covariate adjustments used in the GLGC paper are consistent with those incorporated in our discovery analysis of fatty acid traits We have now added to the manuscript information regarding covariate adjustments for each of the replication and validation analyses (p. 23).
Regarding the sample sizes used for replication and validation, we agree with the reviewer that these analyses exhibit differences in sample size compared to our discovery analyses. Indeed, we chose the GLGC for our validation effort as it is the largest GWAS of lipid levels to date, and we expected it to be well-powered to demonstrate associations with the selected phenotypes. We now include text explaining the motivation of our selection of replication and validation cohorts in the Discusssion (pp. 15-16).